This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. This project aims to investigate glycoconjugate structures of Yersinia pestis and Yersinia pseudotuberculosis biofilms. Biofilms are formed by most pathogenic bacteria. However, no model existed where a living host served as the biofilm target until recently. A biofilm model system is under development where Caenorhabditis elegans is infected with the above bacteria. Mutants in biofilm formation have been generated in Y. pseudotuberculosis and experiments are ongoing to generate biofilm mutants in Yersinia pestis. In bubonic plague the Y. pestis block the flea digestive tract, stimulating the insects to bite humans or other mammals repeatedly in attempts to feed. We have found that Y. pestis also blocks C. elegans from feeding. The blockage is caused by a biofilm, a polysaccharide-rich matrix synthesized by Y. pestis, and requires the bacterial genes hmsHFRS. Significantly, these genes are also required for plague bacteria to block fleas, indicating that Y. pestis uses some of the same pathways to disrupt feeding of both invertebrates. In principle, the study of the biofilm structure in wild-type and mutants will reveal structural detail and aid in the identification of the genes responsible for biofilm polysaccharide biosynthesis and its control. Preliminary study of biofilms from both Yersinia species are under way. GC-MS analysis has revealed the presence of Fuc, Xyl, Man, Glc and trace amounts of Sia, and GlcNAc. Results from MALDI-TOF MS analyses suggest the presence of homopolymers of Hex and heteropolymers containing Hex, HexNAc, deoxyHex and Pent. We are currently developing methods for systematic fractionation and analysis of the biofilm components.